scholarly journals Identification of TGFβ signalling as a regulator of interneuron neurogenesis in a human pluripotent stem cell model

2021 ◽  
Author(s):  
Maria Santos Cruz ◽  
Meng Li
Keyword(s):  
Stem Cells ◽  
Pluripotent Stem Cells ◽  
Cell Model ◽  
Neurological Diseases ◽  
Human Pluripotent Stem Cells ◽  
Tgfβ Signaling ◽  
Neuropsychiatric Diseases ◽  
Cortical Interneurons ◽  
Complex Development ◽  
Tgfβ Signalling

Cortical interneurons are GABAergic inhibitory cells that connect locally in the neocortex and play a  pivotal role in shaping cortical network activities. Dysfunction of these cells is believed to lead to runaway excitation underlying seizure-based diseases, such as epilepsy, autism, and schizophrenia. There is a growing interest in using cortical interneurons derived from human pluripotent stem cells for understanding their complex development and for modeling neuropsychiatric diseases. Here, we report the identification of a novel role of TGFβ signaling in modulating interneuron progenitor maintenance and neuronal differentiation. TGFβ signaling inhibition suppresses terminal differentiation of interneuron progenitors while exogenous TGFβ3 accelerates the transition of progenitors into postmitotic neurons. We provide evidence that TGFb signaling exerts this function via regulating cell cycle length of the NKX2.1+ neural progenitors. Together, this study represents a useful platform for studying human interneuron development and interneuron associated neurological diseases with human pluripotent stem cells.

scholarly journals A Novel Vitronectin Peptide Facilitates Differentiation of Oligodendrocytes from Human Pluripotent Stem Cells (Synthetic ECM for Oligodendrocyte Differentiation)

Biology ◽  
2021 ◽  
Vol 10 (12) ◽  
pp. 1254
Author(s):  
Won Ung Park ◽  
Gyu-Bum Yeon ◽  
Myeong-Sang Yu ◽  
Hui-Gwan Goo ◽  
Su-Hee Hwang ◽  
...  
Keyword(s):  
Stem Cells ◽  
Regenerative Medicine ◽  
Pluripotent Stem Cells ◽  
Neurological Diseases ◽  
Docking Simulation ◽  
Human Pluripotent Stem Cells ◽  
Oligodendrocyte Differentiation ◽  
In Silico Docking ◽  
Cytoskeleton Remodeling ◽  
Culture Environment

Differentiation of oligodendrocytes (ODs) presents a challenge in regenerative medicine due to their role in various neurological diseases associated with dysmyelination and demyelination. Here, we designed a peptide derived from vitronectin (VN) using in silico docking simulation and examined its use as a synthetic substrate to support the differentiation of ODs derived from human pluripotent stem cells. The designed peptide, named VNP2, promoted OD differentiation induced by the overexpression of SOX10 in OD precursor cells compared with Matrigel and full-length VN. ODs differentiated on VNP2 exhibited greater contact with axon-mimicking nanofibers than those differentiated on Matrigel. Transcriptomic analysis revealed that the genes associated with morphogenesis, cytoskeleton remodeling, and OD differentiation were upregulated in cells grown on VNP2 compared with cells grown on Matrigel. This new synthetic VN-derived peptide can be used to develop a culture environment for efficient OD differentiation.

scholarly journals NODAL/TGFβ signalling mediates the self-sustained stemness induced by PIK3CAH1047R homozygosity in pluripotent stem cells

2019 ◽  
Author(s):  
Ralitsa R. Madsen ◽  
James Longden ◽  
Rachel G. Knox ◽  
Xavier Robin ◽  
Franziska Völlmy ◽  
...  
Keyword(s):  
Stem Cells ◽  
Pluripotent Stem Cells ◽  
Human Cancer ◽  
Human Pluripotent Stem Cells ◽  
Depth Analysis ◽  
Pi3k Activation ◽  
Tgfβ Signalling ◽  
Pi3k Signalling ◽  
Pathway Inhibition ◽  
Dose Dependent

AbstractActivating PIK3CA mutations are known “drivers” of human cancer and developmental overgrowth syndromes. We recently demonstrated that the “hotspot” PIK3CAH1047R variant exerts unexpected allele dose-dependent effects on stemness in human pluripotent stem cells (hPSCs). In the present study, we combine high-depth transcriptomics, total proteomics and reverse-phase protein arrays to reveal potentially disease-related alterations in heterozygous cells, and to assess the contribution of activated TGFβ signalling to the stemness phenotype of PIK3CAH1047R homozygous cells. We demonstrate signalling rewiring as a function of oncogenic PI3K signalling dose, and provide experimental evidence that self-sustained stemness is causally related to enhanced autocrine NODAL/TGFβ signalling. A significant transcriptomic signature of TGFβ pathway activation in PIK3CAH1047R heterozygous was observed but was modest and was not associated with the stemness phenotype seen in homozygous mutants. Notably, the stemness gene expression in PIK3CAH1047R homozygous iPSCs was reversed by pharmacological inhibition of TGFβ signalling, but not by pharmacological PI3Kα pathway inhibition. Altogether, this provides the first in-depth analysis of PI3K signalling in human pluripotent stem cells and directly links dose-dependent PI3K activation to developmental NODAL/TGFβ signalling.

scholarly journals A role for TGFβ signalling in medium spiny neuron differentiation of human pluripotent stem cells

2020 ◽  
Vol 4 (2) ◽  
Author(s):  
Marija Fjodorova ◽  
Zoe Noakes ◽  
Meng Li
Keyword(s):  
Stem Cells ◽  
Pluripotent Stem Cells ◽  
Activin A ◽  
Human Pluripotent Stem Cells ◽  
Medium Spiny Neuron ◽  
Fine Tuning ◽  
Fate Specification ◽  
Differentiation Protocol ◽  
Tgfβ Signalling ◽  
Bmp Signalling

Abstract Activin A and other TGFβ family members have been shown to exhibit a certain degree of promiscuity between their family of receptors. We previously developed an efficient differentiation protocol using Activin A to obtain medium spiny neurons (MSNs) from human pluripotent stem cells (hPSCs). However, the mechanism underlying Activin A-induced MSN fate specification remains largely unknown. Here we begin to tease apart the different components of TGFβ pathways involved in MSN differentiation and demonstrate that Activin A acts exclusively via ALK4/5 receptors to induce MSN progenitor fate during differentiation. Moreover, we show that Alantolactone, an indirect activator of SMAD2/3 signalling, offers an alternative approach to differentiate hPSC-derived forebrain progenitors into MSNs. Further fine tuning of TGFβ pathway by inhibiting BMP signalling with LDN193189 achieves accelerated MSN fate specification. The present study therefore establishes an essential role for TGFβ signalling in human MSN differentiation and provides a fully defined and highly adaptable small molecule-based protocol to obtain MSNs from hPSCs.

scholarly journals NODAL/TGFβ signalling mediates the self-sustained stemness induced by PIK3CAH1047R homozygosity in pluripotent stem cells

2021 ◽  
pp. dmm.048298
Author(s):  
Ralitsa R. Madsen ◽  
James Longden ◽  
Rachel G. Knox ◽  
Xavier Robin ◽  
Franziska Völlmy ◽  
...  
Keyword(s):  
Stem Cells ◽  
Pluripotent Stem Cells ◽  
Human Cancer ◽  
Human Pluripotent Stem Cells ◽  
Human Genetic Disease ◽  
Allele Dosage ◽  
Depth Analysis ◽  
Tgfβ Signalling ◽  
Pi3k Signalling ◽  
Pathway Inhibition

Activating PIK3CA mutations are known “drivers” of human cancer and developmental overgrowth syndromes. We recently demonstrated that the "hotspot" PIK3CAH1047R variant exerts unexpected allele dose-dependent effects on stemness in human pluripotent stem cells (hPSCs). In the present study, we combine high-depth transcriptomics, total proteomics and reverse-phase protein arrays to reveal potentially disease-related alterations in heterozygous cells, and to assess the contribution of activated TGFβ signalling to the stemness phenotype of homozygous PIK3CAH1047R cells. We demonstrate signalling rewiring as a function of oncogenic PI3K signalling strength, and provide experimental evidence that self-sustained stemness is causally related to enhanced autocrine NODAL/TGFβ signalling. A significant transcriptomic signature of TGFβ pathway activation in heterozygous PIK3CAH1047R was observed but was modest and was not associated with the stemness phenotype seen in homozygous mutants. Notably, the stemness gene expression in homozygous PIK3CAH1047R iPSCs was reversed by pharmacological inhibition of NODAL/TGFβ signalling, but not by pharmacological PI3Kα pathway inhibition. Altogether, this provides the first in-depth analysis of PI3K signalling in human pluripotent stem cells and directly links strong PI3K activation to developmental NODAL/TGFβ signalling. This work illustrates the importance of allele dosage and expression when artificial systems are used to model human genetic disease caused by activating PIK3CA mutations.

scholarly journals Generating microglia from human pluripotent stem cells: novel in vitro models for the study of neurodegeneration

2019 ◽  
Vol 14 (1) ◽  
Author(s):  
Anna M. Speicher ◽  
Heinz Wiendl ◽  
Sven G. Meuth ◽  
Matthias Pawlowski
Keyword(s):  
Stem Cells ◽  
Pluripotent Stem Cells ◽  
Drug Testing ◽  
Disease Modeling ◽  
Current Knowledge ◽  
Neurological Diseases ◽  
Human Pluripotent Stem Cells ◽  
Comprehensive Overview ◽  
Human Microglia

AbstractMicroglia play an essential role for central nervous system (CNS) development and homeostasis and have been implicated in the onset, progression, and clearance of numerous diseases affecting the CNS. Previous in vitro research on human microglia was restricted to post-mortem brain tissue-derived microglia, with limited availability and lack of scalability. Recently, the first protocols for the generation of microglia from human pluripotent stem cells have become available, thus enabling the implementation of powerful platforms for disease modeling, drug testing, and studies on cell transplantation. Here we give a detailed and comprehensive overview of the protocols available for generating microglia from human pluripotent stem cells, highlighting the advantages, drawbacks, and operability and placing them into the context of current knowledge of human embryonic development. We review novel insights into microglia biology and the role of microglia in neurological diseases as drawn from the new methods and provide an outlook for future lines of research involving human pluripotent stem cell-derived microglia.

scholarly journals Global Transcriptional Analyses of the Wnt-Induced Development of Neural Stem Cells from Human Pluripotent Stem Cells

2021 ◽  
Vol 22 (14) ◽  
pp. 7473
Author(s):  
Bing-Chun Liu ◽  
Fang-Yuan Liu ◽  
Xin-Yue Gao ◽  
Yang-Lin Chen ◽  
Qiao-Qiao Meng ◽  
...  
Keyword(s):  
Stem Cells ◽  
Neural Stem Cells ◽  
Cell Fate ◽  
Pluripotent Stem Cells ◽  
Hedgehog Signaling ◽  
Cell Model ◽  
Human Pluripotent Stem Cells ◽  
Cell Fate Decisions ◽  
Neural Lineage

The differentiation of human pluripotent stem cells (hPSCs) to neural stem cells (NSCs) is the key initial event in neurogenesis and is thought to be dependent on the family of Wnt growth factors, their receptors and signaling proteins. The delineation of the transcriptional pathways that mediate Wnt-induced hPSCs to NSCs differentiation is vital for understanding the global genomic mechanisms of the development of NSCs and, potentially, the creation of new protocols in regenerative medicine. To understand the genomic mechanism of Wnt signaling during NSCs development, we treated hPSCs with Wnt activator (CHIR-99021) and leukemia inhibitory factor (LIF) in a chemically defined medium (N2B27) to induce NSCs, referred to as CLNSCs. The CLNSCs were subcultured for more than 40 passages in vitro; were positive for AP staining; expressed neural progenitor markers such as NESTIN, PAX6, SOX2, and SOX1; and were able to differentiate into three neural lineage cells: neurons, astrocytes, and oligodendrocytes in vitro. Our transcriptome analyses revealed that the Wnt and Hedgehog signaling pathways regulate hPSCs cell fate decisions for neural lineages and maintain the self-renewal of CLNSCs. One interesting network could be the deregulation of the Wnt/β-catenin signaling pathway in CLNSCs via the downregulation of c-MYC, which may promote exit from pluripotency and neural differentiation. The Wnt-induced spinal markers HOXA1-4, HOXA7, HOXB1-4, and HOXC4 were increased, however, the brain markers FOXG1 and OTX2, were absent in the CLNSCs, indicating that CLNSCs have partial spinal cord properties. Finally, a CLNSC simple culture condition, when applied to hPSCs, supports the generation of NSCs, and provides a new and efficient cell model with which to untangle the mechanisms during neurogenesis.

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